Developmental regulation of the human cathepsin G gene in myelomonocytic cells.
Cathepsin G is a neutral serine protease that is found in the azurophil granules of neutrophils and monocytes. Previous experiments had demonstrated that cathepsin G is actively produced by the promonocytic U937 cell line, and that 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced differentiation of these cells toward macrophages resulted in a reduction of cathepsin G activity. In this study, we have analyzed the mechanism of this TPA-induced down-regulatory event. Using in situ hybridization techniques, we demonstrated that cathepsin G mRNA is detectable only at the promyelocytic stage of myeloid development. Using U937 promonocytic cells as a model, we demonstrated; 1) cathepsin G protein levels decline in TPA-treated cells; 2) this decline was due to a nearly complete loss of cathepsin G mRNA in cells treated with TPA for 24 h; and 3) the rate of cathepsin G mRNA loss with TPA treatment was similar to that with actinomycin D. These results suggested that cathepsin G transcription was down-regulated within several hours of TPA addition. This was directly tested by performing nuclear run-off assays of TPA-treated U937 cells; cathepsin G transcription was shown to be strand-specific, and declined within 4 h of TPA addition. Cathepsin G transcription was essentially undetectable 8 or more hours after TPA treatment, suggesting that down-regulation is predominantly transcriptional. Cycloheximide treatment of U937 cells resulted in a partial block of TPA-mediated cathepsin G down-regulation, indicating that continuous protein synthesis is required for down-regulation to occur. A newly synthesized protein or proteins may therefore be required for the transcriptional down-regulation of cathepsin G during the normal development of promyelocytes or promonocytes.
Hanson
R D
RD
Department of Medicine Jewish Hospital, Washington University Medical Center, St. Louis, Missouri 63110.
Connolly
N L
NL
Burnett
D
D
Campbell
E J
EJ
Senior
R M
RM
Ley
T J
TJ
D056649:Cathepsin G; D002403:Cathepsins / Q000235:genetics* / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D009000:Monocytes / Q000502:physiology*; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000235:genetics; D012697:Serine Endopeptidases; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects
D012333:RNA, Messenger; D003609:Dactinomycin; D003513:Cycloheximide; D002403:Cathepsins; D012697:Serine Endopeptidases; C534094:CTSG protein, human; D056649:Cathepsin G; D013755:Tetradecanoylphorbol Acetate